Many methods have been developed for the qualitative and quantitative determination of low molecular weight substances such as hormones and the like. Amongst the most sensitive are those based on proteins capable of binding specifically the substance to be determined. A very sensitive test is the radio-immunoassay in its variations. Radio immunoassays have the drawback that the labelled entity has a comparatively short shelf-life, that there exist severe regulations concerning the handling of radioactive materials and that highly qualified personnel must be used for performing such assays. A multitude of Enzyme-Immunoassays is known, and part of these is summarized in the review of this subject in Clinical Chemistry 22 (1976) 1243-1255.
The present invention provides a sensitive and convenient quantitative assay for the determination of small molecules (haptens) such as hormones, vitamins, glycosides, etc. Haptens are defined as protein-free substances whose chemical configuration is such that they can react with specific antibodies but are themselves not capable of causing the formation of antibodies. In order to produce antibodies against specific haptens, such haptens must be first coupled to macromolecules and the resulting conjugate is injected into suitable test animals.